Figure 6.

ATM-Dependent Phosphorylation of 53BP1 Promotes RIF1 Interaction

(A) Irradiated WT MEFs pretreated with DMSO or ATMi were fixed and immunostained for γH2AX and either RIF1 or 53BP1.

(B) Quantification of 53BP1 and RIF1 IRIF in cells prepared as in (A). n > 4 ± SD, >126 cells scored per condition.

(C) Immunoblots of whole-cell lysates prepared from 53Bp1^−/− MEFs complemented with the indicated constructs as in Figure 5 1 hr following mock or 10 Gy IR.

(D) Flag-HA-53BP1 proteins were purified with Flag-M2 beads (Sigma) from lysates in (C) under stringent conditions. Bead complexes were subsequently incubated in nuclear extracts (HNE) before 53BP1-associated proteins were recovered by peptide elution. B/O = beads-only control.

(E) Immunoblots of whole-cell lysates of 53Bp1^−/− MEFs complemented with WT 53BP1, 1 hr following mock or 15 Gy IR treatments in the presence/absence of ATMi.

(F) As in (D), but with FLAG-HA-53BP1 purified from lysates indicated in (E). See also Figure S6.